Several lines of evidence suggest that mast cells promote tumor angiogenesis. The long-term objective of this proposal is to understand the molecular interactions between mast cells and endothelial cells in the development of tumor angiogenesis, in particular the role of ephrinB2 and EphB4 in this process. EphB4 is one member of a receptor tyrosine kinase family, and its co-receptor is ephrinB2. These transmembrane proteins require cell-cell contact for activation and are essential in vascular development and angiogenesis. We have shown that mast cells are juxtaposed to basement membranes of angiogenic capillaries of dysplastic skin in a mouse model of skin squamous cell carcinoma and that mast cells are involved in the "angiogenic switch," in which angiogenesis is turned on at early stages of neoplastic development. Therefore, we hypothesize that Eph-ephrin signaling between mast cells and endothelial cells is essential in activation of angiogenesis and progression to tumor development. We propose to pursue several specific aims as follows: AIM 1: Determine whether mast cell EphB4 and ephrinB2 are essential in murine models of epithelial tumor angiogenesis; AIM 2: Determine whether mast cell-endothelial cell interactions are regulated by EphB4 and ephrinB2 signaling; AIM 3: Determine whether mast cell function is regulated by EphB4 and ephrinB2 signaling. To achieve these aims we plan to use two mouse models of carcinoma, a skin squamous cell carcinoma (K14-HPV16) and a bronchoalveolar carcinoma (CC10-SV40Tag) model. We will also utilize mice whose tissues can be stained with LacZ as a surrogate for ephrinB2 and EphB4 expression, and mice that are mast cell-deficient (KitWsh/KitWsh). In vitro studies will use knockout mouse embryonic stem cells that will be differentiated into mast cells in culture and later used to reconstitute tissues of transgenic cancer mice. These studies will further the understanding of molecular mechanisms behind cancer progression and elucidate specific targets opposing malignant progression by interfering with the angiogenic switch. [unreadable] [unreadable]